Methods using 3-(4-amino-1-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione for treatment of certain leukemias

ABSTRACT

Methods of treating, preventing or managing leukemias are disclosed. The methods encompass the administration of an immunomodulatory compound of the invention known as Revlimid® or lenalidomide. The invention further relates to methods of treatment using this compound with chemotherapy, radiation therapy, hormonal therapy, biological therapy or immunotherapy. Pharmaceutical compositions and single unit dosage forms suitable for use in the methods of the invention are also disclosed.

This application is a continuation-in-part of U.S. patent applicationSer. No. 10/438,213, filed May 15, 2003, which claims the benefit ofU.S. provisional application Nos. 60/380,842, filed May 17, 2002, and60/424,600, filed Nov. 6, 2002, the entireties of which are incorporatedherein by reference.

1. FIELD OF THE INVENTION

This invention relates to methods of treating, preventing or managingleukemias with an immunomodulatory compound having the chemical name of3-(4-amino-1-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione, whichis also known as Revlimid® or Revimid®. In particular, this inventionencompasses methods of treating, preventing or managing leukemias,including but not limited to, chronic lymphocytic leukemia, chronicmyelocytic leukemia, acute lymphoblastic leukemia, acute myelogenousleukemia and acute myeloblastic leukemia, using the compound alone as atherapeutic.

The invention also encompasses the use of specific combinations or“cocktails” of Revlimid® and other therapy, e.g., radiation or otherchemotherapeutics, including but not limited to, anti-cancer agents,immunosuppressive agents, and anti-inflammatories such as steroids. Theinvention also relates to pharmaceutical compositions and dosingregimens with said compound alone that is as a therapeutic.

2. BACKGROUND OF THE INVENTION

2.1 Pathobiology of Cancer

Cancer is characterized primarily by an increase in the number ofabnormal cells derived from a given normal tissue, invasion of adjacenttissues by these abnormal cells, or lymphatic or blood-borne spread ofmalignant cells to regional lymph nodes and to distant sites(metastasis). Clinical data and molecular biologic studies indicate thatcancer is a multistep process that begins with minor preneoplasticchanges, which may under certain conditions progress to neoplasia. Theneoplastic lesion may evolve clonally and develop an increasing capacityfor invasion, growth, metastasis, and heterogeneity, especially undercondition in which the neoplastic cells escape the host's immunesurveillance. Roitt, I., Brostoff, J. and Kale, D., Immunology,17.1-17.12 (3rd ed., Mosby, St. Louis, Mo., 1993).

There is an enormous variety of cancers which are described in detail inthe medical literature. Examples includes cancer of the blood, lung,colon, rectum, prostate, breast, brain, and intestine. The various formsof the cancers such as leukemias are described in U.S. provisionalapplication No. 60/380,842, filed May 17, 2002, the entireties of whichare incorporated herein by reference (see, e.g., Section 2.2. Types ofCancers).

In particular, leukemia refers to malignant neoplasms of theblood-forming tissues. Although viruses reportedly cause several formsof leukemia in animals, causes of leukemia in humans are to a largeextend unknown. The Merck Manual, 944-952 (17^(th) ed. 1999).Transformation to malignancy typically occurs in a single cell throughtwo or more steps with subsequent proliferation and clonal expansion. Insome leukemias, specific chromosomal translocations have been identifiedwith consistent leukemic cell morphology and special clinical features(e.g., translocations of 9 and 22 in chronic myelocytic leukemia, and of15 and 17 in acute promyelocytic leukemia). Acute leukemias arepredominantly undifferentiated cell populations and chronic leukemiasmore mature cell forms.

Acute leukemias are divided into lymphoblastic (ALL) andnon-lymphoblastic (ANLL) types. The Merck Manual, 946-949 (17^(th) ed.1999). They may be further subdivided by their morphologic andcytochemical appearance according to the French-American-British (FAB)classification or according to their type and degree of differentiation.The use of specific B- and T-cell and myeloid-antigen monoclonalantibodies are most helpful for classification. ALL is predominantly achildhood disease which is established by laboratory findings and bonemarrow examination. ANLL, also known as acute myelogenous leukemia oracute myeloblastic leukemia (AML), occurs at all ages and is the morecommon acute leukemia among adults; it is the form usually associatedwith irradiation as a causative agent.

Chronic leukemias are described as the lymphocytic (CLL) or myelocytic(CML). The Merck Manual, 949-952 (17^(th) ed. 1999). CLL ischaracterized by the appearance of mature lymphocytes in blood, bonemarrow, and lymphoid organs. The hallmark of CLL is sustained, absolutelymphocytosis (>5,000/μL) and an increase of lymphocytes in the bonemarrow. Most CLL patients also have clonal expansion of lymphocytes withB-cell characteristics. CLL is a disease of middle or old age. In CML,the characteristic feature is the predominance of granulocytic c3ells ofall stages of differentiation in blood, bone marrow, liver, spleen, andother organs. In the sympotomatic patient at diagnosis, the total WBCcount is usually about 200,000/μL, but may reach 1,000,000/μL. CML isrelatively easy to diagnose because of the presences of the Philadelphiachromosome.

The incidence of cancer continues to climb as the general populationages, as new cancers develop, and as susceptible populations (e.g.,people infected with AIDS or excessively exposed to sunlight) grow. Inparticular, chronic lymphocytic leukemia is an incurable leukemia withlimited therapeutic options for patients with relapsed or refractorydisease. A tremendous demand therefore exists for new methods andcompositions that can be used to treat patients with cancer includingleukemia.

Many types of cancers are associated with new blood vessel formation, aprocess known as angiogenesis. Several of the mechanisms involved intumor-induced angiogenesis have been elucidated. The most direct ofthese mechanisms is the secretion by the tumor cells of cytokines andangiogenic properties. Examples of these cytokines include acidic andbasic fibroblastic growth factor (a,b-FGF), angiogenin, vascularendothelial growth factor (VEGF), and TNF-α. Alternatively, tumor cellscan release angiogenic peptides through the production of proteases andthe subsequent breakdown of the extracellular matrix where somecytokines are stored (e.g., b-FGF). Angiogensis can also be inducedindirectly through the recruitment of inflammatory cells (particularlymacrophages) and their subsequent release of angiogenic cytokines (e.g.,TNF-α, bFGF).

Accordingly, compounds that can control angiogenesis or inhibit theproduction of certain cytokines, including TNF-α, may be useful in thetreatment and prevention of various cancers.

2.2 Methods of Treating Cancer

Current cancer therapy may involve surgery, chemotherapy, hormonaltherapy and/or radiation treatment to eradicate neoplastic cells in apatient (see, for example, Stockdale, 1998, Medicine, vol. 3, Rubensteinand Federman, eds., Chapter 12, Section IV). Recently, cancer therapycould also involve biological therapy or immunotherapy. All of theseapproaches pose significant drawbacks for the patient. Surgery, forexample, may be contraindicated due to the health of a patient of may beunacceptable to the patient. Additionally, surgery may not completelyremove neoplastic tissue. Radiation therapy is only effective when theneoplastic tissue exhibits a higher sensitivity to radiation than normaltissue. Radiation therapy can also often elicit serious side effects.Hormonal therapy is rarely given as a single agent. Although hormonaltherapy can be effective, it is often used to prevent or delayrecurrence of cancer after other treatments have removed the majority ofcancer cells. Biological therapies and immunotherapies are limited innumber and may produce side effects such as rashes or swellings,flu-like symptoms, including fever, chills and fatigue, digestive tractproblems or allergic reactions.

With respect to chemotherapy, there are a variety of chemotherapeuticagents available for treatment of cancer. A majority of cancerchemotherapeutics act by inhibiting DNA synthesis, either directly, orindirectly by inhibiting the biosynthesis of deoxyribonucleotidetriphosphate precursors, to prevent DNA replication and concomitant celldivision. Gilman et al., Goodman and Gilman's: The Pharmacological Basisof Therapeutics, Tenth Ed. (McGraw Hill, New York).

Despite availability of a variety of chemotherapeutic agents,chemotherapy has many drawbacks. Stockdale, Medicine, vol. 3, Rubensteinand Federman, eds., ch. 12, sect. 10, 1998. Almost all chemotherapeuticagents are toxic, and chemotherapy causes significant, and oftendangerous side effects including severe nausea, bone marrow depression,and immunosuppression. Additionally, even with administration ofcombinations of chemotherapeutic agents, many tumor cells are resistantor develop resistance to the chemotherapeutic agents. In fact, thosecells resistant to the particular chemotherapeutic agents used in thetreatment protocol often prove to be resistant to other drugs, even ifthose agents act by different mechanism from those of the drugs used inthe specific treatment. This phenomenon is referred to as pleiotropicdrug or multidrug resistance. Because of the drug resistance, manycancers prove refractory to standard chemotherapeutic treatmentprotocols.

Still, there is a significant need for safe and effective methods oftreating, preventing and managing cancer, particularly for diseases thatare refractory to standard treatments, such as surgery, radiationtherapy, chemotherapy and hormonal therapy, while reducing or avoidingthe toxicities and/or side effects associated with the conventionaltherapies.

2.3 IMiDs®

A number of studies have been conducted with the aim of providingcompounds that can safely and effectively be used to treat diseasesassociated with abnormal production of TNF-α. See, e.g., Marriott, J.B., et al., Expert Opin. Biol. Ther. 1(4):1-8 (2001); G. W. Muller, etal., Journal of Medicinal Chemistry 39(17): 3238-3240 (1996); and G. W.Muller, et al., Bioorganic & Medicinal Chemistry Letters 8: 2669-2674(1998). Some studies have focused on a group of compounds selected fortheir capacity to potently inhibit TNF-α production by LPS stimulatedPBMC. L. G. Corral, et al., Ann. Rheum. Dis. 58:(Suppl 1) 1107-1113(1999). These compounds, which are referred to as IMiDs®(CelgeneCorporation) or Immunomodulatory Drugs, show not only potent inhibitionof TNF-α but also inhibition of LPS induced monocyte IL1β and IL12production. LPS induced IL6 is also inhibited by immunomodulatorycompounds of the invention, albeit partially. These compounds are potentstimulators of LPS induced IL10, Id. Particular examples of IMiDs®include, but are not limited to, the substituted2-(2,6-dioxopiperidin-3-yl)phthalimides and substituted2-(2,6-dioxopiperidin-3-yl)-1-oxoisoindoles described in U.S. Pat. Nos.5,635,517, 6,281,230 and 6,316,471, to G. W. Muller, et al.

3. SUMMARY OF THE INVENTION

This invention encompasses methods of treating, preventing or managingcertain types of cancer, including primary and metastic cancer, as wellas cancers that are relapsed, refractory or resistant to conventionalchemotherapy. In particular, methods of this invention encompass methodsof treating, preventing or managing various forms of leukemias such aschronic lymphocytic leukemia, chronic myelocytic leukemia, acutelymphoblastic leukemia, acute myelogenous leukemia and acutemyeloblastic leukemia, including leukemias that are relapsed, refractoryor resistant.

The methods comprise administering to a patient in need of suchtreatment, prevention or management a therapeutically orprophylactically effective amount of an immunomodulatory compound of theinvention, or a pharmaceutically acceptable salt, solvate, hydrate,stereoisomer, clathrate, or prodrug thereof. In a preferred embodiment,the immunomodulatory compound is used alone, that is without otherchemotherapeutics.

In another methods of the invention, an immunomodulatory compound of theinvention is administered in combination with a therapy conventionallyused to treat, prevent or manage cancer. Examples of such conventionaltherapies include, but are not limited to, surgery, chemotherapy,radiation therapy, hormonal therapy, biological therapy, immunotherapyand combinations thereof.

This invention also encompasses pharmaceutical compositions, single unitdosage forms, and dosing regimens which comprise an immunomodulatorycompound of the invention, or a pharmaceutically acceptable salt,solvate, hydrate, stereoisomer, clathrate, or prodrug thereof, and asecond, or additional, active agent. Second active agents includespecific combinations, or “cocktails,” of drugs or therapy, or both.

The preferred compound to be used in the methods and composition is3-(4-amino-1-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione(Revlimid®).

4. BRIEF DESCRIPTION OF FIGURE

FIG. 1 shows a comparison of the effects of3-(4-amino-1-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione(Revlimid®) and thalidomide in inhibiting the proliferation of multiplemyelome (MM) cell lines in an in vitro study. The uptake of[3H]-thymidine by different MM cell lines (MM, 1S, Hs Sultan, U266 andRPMI-8226) was measured as an indicator of the cell proliferation.

5. DETAILED DESCRIPTION OF THE INVENTION

A first embodiment of the invention encompasses methods of treating,managing, or preventing cancer which comprises administering to apatient in need of such treatment, management or prevention atherapeutically or prophylactically effective amount of animmunomodulatory compound of the invention, or a pharmaceuticallyacceptable salt, solvate, hydrate, stereoisomer, clathrate, or prodrugthereof. In particular, methods of this invention encompass methods oftreating, preventing or managing various forms of leukemias, includingbut not limited to, chronic lymphocytic leukemia, chronic myelocyticleukemia, acute lymphoblastic leukemia, acute myelogenous leukemia andacute myeloblastic leukemia. In one embodiment, the leukemia isrefractory leukemia, relapsed leukemia or a leukemia that is resistantto chemotherapy other than an immunomodulatory compound of theinvention.

In a separate and distinct embodiment of the invention, theimmunomodulatory compound of the invention is administered incombination with another drug (“second active agent”) or another therapyfor treating, managing, or preventing cancer. Second active agentsinclude small molecules and large molecules (e.g., proteins andantibodies), examples of which are provided herein, as well as stemcells or cord blood. Methods, or therapies, that can be used incombination with the administration of an immunomodulatory compound ofthe invention include, but are not limited to, surgery, bloodtransfusions, immunotherapy, biological therapy, radiation therapy, andother non-drug based therapies presently used to treat, prevent ormanage cancer.

The invention also encompasses pharmaceutical compositions (e.g., singleunit dosage forms) that can be used in methods disclosed herein.Particular pharmaceutical compositions comprise an immunomodulatorycompound of the invention, or a pharmaceutically acceptable salt,solvate, hydrate, stereoisomer, clathrate, or prodrug thereof, and asecond active agent.

5.1 Immunomodulatory Compounds

Compounds used in the invention include compounds that are racemic,stereomerically enriched or stereomerically pure. In some embodiments,pharmaceutically acceptable salts, solvates, hydrates, clathrates, andprodrugs thereof are included. Preferred compounds used in the inventionare small organic molecules having a molecular weight less than about1,000 g/mol, and are not proteins, peptides, oligonucleotides,oligosaccharides or other macromolecules.

As used herein and unless otherwise indicated, the terms“immunomodulatory compounds” and “IMiDs®” (Celgene Corporation)encompasses small organic molecules that markedly inhibit TNF-α, LPSinduced monocyte IL1β and IL12, and partially inhibit IL6 production.Specific immunomodulatory compounds of the invention are discussedbelow.

In the most preferred embodiment, “an immunomodulatory compound of theinvention” refers to3-(4-amino-1-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione(lenalidomide, also known as Revlimid® or Revimid®). The compound3-(4-amino-1-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione has thefollowing chemical structure:

Specific examples of immunomodulatory compounds, include, but are notlimited to, cyano and carboxy derivatives of substituted styrenes suchas those disclosed in U.S. Pat. No. 5,929,117;1-oxo-2-(2,6-dioxo-3-fluoropiperidin-3yl) isoindolines and1,3-dioxo-2-(2,6-dioxo-3-fluoropiperidine-3-yl) isoindolines such asthose described in U.S. Pat. No. 5,874,448; the tetra substituted2-(2,6-dioxopiperdin-3-yl)-1-oxoisoindolines described in U.S. Pat. No.5,798,368; 1-oxo and 1,3-dioxo-2-(2,6-dioxopiperidin-3-yl) isoindolines,including, but not limited to, those disclosed in U.S. Pat. No.5,635,517; substituted 2-(2,6-dioxopiperidin-3-yl) phthalimides andsubstituted 2-(2,6-dioxopiperidin-3-yl)-1-oxoisoindoles such as thosedescribed in U.S. Pat. Nos. 6,281,230 and 6,316,471; a class ofnon-polypeptide cyclic amides disclosed in U.S. Pat. Nos. 5,698,579 and5,877,200; thalidomide analogs, including hydrolysis products,metabolites, and precursors of thalidomide, such as those described inU.S. Pat. Nos. 5,593,990, 5,629,327, and 6,071,948 to D'Amato; andisoindole-imide compounds such as those described in U.S. patentpublication no. 2003/0096841, and International Application No.PCT/US01/50401 (International Publication No. WO 02/059106). Theentireties of each of the patents and patent applications identifiedherein are incorporated herein by reference. Immunomodulatory compoundsof the invention do not include thalidomide.

The immunomodulatory compounds of the invention can either becommercially purchase or prepared according to the methods described inthe patents or pate publication disclosed herein (see e.g., U.S. Pat.No. 5,635,517, incorporated herein by reference). Further, opticallypure compounds can be asymmetrically synthesized or resolved using knownresolving agents or chiral columns as well as other standard syntheticorganic chemistry techniques.

As used herein and unless otherwise indicated, the term“pharmaceutically acceptable salt” encompasses non-toxic acid and baseaddition salts of the compound to which the term refers. Acceptablenon-toxic acid addition salts include those derived from organic andinorganic acids or bases know in the art, which include, for example,hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid,methanesulphonic acid, acetic acid, tartaric acid, lactic acid, succinicacid, citric acid, malic acid, maleic acid, sorbic acid, aconitic acid,salicylic acid, phthalic acid, embolic acid, enanthic acid, and thelike.

Compounds that are acidic in nature are capable of forming salts withvarious pharmaceutically acceptable bases. The bases that can be used toprepare pharmaceutically acceptable bases. The bases that can be used toprepare pharmaceutically acceptable base addition salts of such acidiccompounds are those that form non-toxic base addition salts, i.e., saltscontaining pharmacologically acceptable cations such as, but not limitedto, alkali metal or alkaline earth metal salts and the calcium,magnesium, sodium or potassium salts in particular. Suitable organicbases include, but are not limited to, N,N-dibenzylethylenediamine,chloroprocaine, choline, diethanolamine, ethylenediamine, meglumaine(N-methylglucamine), lysine, and procaine.

As used herein and unless otherwise indicated the term “prodrug” means aderivative of a compound that can hydrolyze, oxidize, or otherwise reactunder biological conditions (in vitro or in vivo) to provide thecompound. Examples of prodrugs include, but are not limited to,derivatives of immunomodulatory compounds of the invention that comprisebiohydrolyzable moieties such as biohydrolyzable amides, biohydrolyzableesters, biohydrolyzable carbamates, biohydrolyzable carbonates,biohydrolyzable ureides, and biohydrolyzable phosphate analogues. Otherexamples of prodrugs include derivatives of immunomodulatory compoundsof the invention that comprise —NO, —NO₂, —ONO, or —ONO₂ moieties.Prodrugs can typically be prepared using well-know methods, such asthose described in I Burger'Medicinal Chemistry and Drug Discovery,172-78, 949-982 (Manfred E. Wolff ed., 5th ed. 1995), and Design ofProdrugs (H. Bundgaard ed., Elselvier, N.Y. 1985).

As used herein and unless otherwise indicated, the terms“biohydrolyzable amide,” “biohydrolyzable ester,” “biohydrolyzablecarbamate,” “biohydrolyzable carbonate,” “biohydrolyzable ureide,”“biohydrolyzable phosphate” mean an amide, ester, carbamate, carbonateureide, or phosphate, respectively, of a compound that either: 1) doesnot interfere with the biological activity of the compound but canconfer upon that compound advantageous properties in vivo, such asuptake, duration of action, or onset of compound advantageous propertiesin vivo, such as uptake, duration of action, or onset of action; or 2)is biologically inactive but is converted in vivo to the biologicallyactive compound. Examples of biohydrolyzable esters include, but are notlimited to, lower alkyl esters, lower acyloxyalkyl esters (such asacetoxylmethyl, acetoxyethyl, aminocarbonyloxymethyl, pivaloyloxymethyl,and pivaloyloxyethyl esters), lactonyl esters (such as phthalidyl andthiophthalidyl esters), lower alkoxyacyloxyalkyl esters (such asmethoxycarbonyl-oxymethyl, ethoxycarbonyloxyethyl andisopropoxycarbonyloxyethyl esters), alkoxyalkyl esters, choline ester,and acylamino alkyl esters (such as acetamidomethyl esters). Examples ofbiohydrolyzable amides include, but are not limited to, lower alkylamides, α-amides acid amides, alkoxyacyl amides, andalkylaminoalkylcarbonyl amides. Examples of biohydrolyzable carbamatesinclude, but are no limited to, lower alkylamines, substitutedethylenediamines, amino acids, hydroxyalkylamines, heterocyclic andheteroaromatic amines, and polyether amines.

The immunomodulatory compound of the invention contains a chiral center,and thus can exist as a racemic mixture of R and S enantiomers. Thisinvention encompasses the use of stereomerically pure forms of thiscompound, as well as the use of mixtures of those forms. For examples,mixtures comprising equal or unequal amounts of the enantiomers may beused in methods and compositions of the invention. These isomers may beasymmetrically synthesized or resolved using standard techniques such aschiral columns or chiral resolving agents. See, e.g., Jacques, J., etal., Enantiomers, Racemates and Resolutions (Wiley-Interscience, NewYork, 1981); Wilen, S. H., et al., Tetrahedron 33:2725 (1977); Eliel, E.L., Stereochemistry of Carbon Compounds (McGraw-Hill, NY, 1926); andWilen, S. H., Tables of Resolving Agents and Optical Resolutions p. 268(E. L. Eliel, Ed., Univ. of Notre Dame Press, Notre Dame, Ind., 1972).

As used herein and unless otherwise indicated, the term “stereomericallypure” means a compositions that comprises one stereoisomer of a compoundand is substantially free of other stereoisomers of that compound. Forexample, a stereomerically pure composition of a compound having onechiral center will be substantially free of the opposite enantiomer ofthe compound. A stereomerically pure composition of a compound havingtwo chiral centers will be substantially free of other diastereomers ofthe compound. A typical stereomerically pure compound comprises greaterthan about 80% by weight of one stereoisomer of the compound and lessthan about 20% by weight of other stereoisomers of the compound, morepreferably greater than about 90% by weight of one stereoisomer of thecompound and less than about 10% by weight of the other stereoisomers ofthe compound, even more preferably greater than about 95% by weight ofone stereoisomer of the compound and less than about 5% by weight of theother stereoisomers of the compound, and most preferably greater thanabout 97% by weight of one stereoisomer of the compound and less thanabout 3% by weight of the other stereoisomers of the compound. As usedherein and unless otherwise indicated, the term “stereomericallyenriched” means a composition that comprises greater than about 60% byweight of one stereoisomer of a compound, preferably greater than about70% by weight, more preferably greater than about 80% by weight of onestereoisomer of a compound. As used herein and unless otherwiseindicated, the term “enantiomerically pure” means a stereomerically purecomposition of a compound having one chiral center. Similarly, the term“stereomerically enriched” means a stereomerically enriched compositionof a compound having one chiral center. In other words, the inventionencompasses the use of the R or S enantiomer of immunomodulatorycompound in the methods.

It should be noted that if there is a discrepancy between a depictedstructure and a name given that structure, the depicted structure is tobe accorded more weight. In addition, if the stereochemistry of astructure of a portion of a structure is not indicated with, forexample, bold or dashed lines, the structure of portion of the structureis to be interpreted as encompassing all stereoisomers of it.

5.2 Second Active Agents

An immunomodulatory compound of the invention can be used with orcombined with other pharmacologically active compounds (“second activeagents”) in methods and compositions of the invention. It is believedthat certain combinations work synergistically in the treatment ofparticular types of cancers, and certain diseases and conditionsassociated with, or characterized by, undesired angiogenesis.Immunomodulatory compounds of the invention can also work to alleviateadverse effects associated with certain second active agents, and somesecond active agents can be used to alleviate adverse effects associatedwith immunomodulatory compounds of the invention.

One or more second active ingredients or agents can be used in themethods and compositions of the invention together with animmunomodulatory compound of the invention. Second active agents can belarge molecules (e.g., proteins) or small molecules (e.g., syntheticinorganic, organometallic, or organic molecules).

Examples of large molecule active agents include, but are not limitedto, hematopoietic growth factors, cytokines, and monoclonal andpolyclonal antibodies. Typical large molecule active agents arebiological molecules, such as naturally occurring or artificially madeproteins. Proteins that are particularly useful in this inventioninclude proteins that stimulate the survival and/or proliferation ofhematopoietic precursor cells and immunologically active poietic cellsin vitro or in vivo. Others stimulate the division and differentiationof committed erythroid progenitors in cells in vitro or in vivo.Particular proteins include, but are not limited to: interleukins, suchas IL-2 (including recombinant IL-11 (“rIL2”) and canarypox IL-2),IL-10, IL-12, and IL-18; interferons, such as interferon alfa-2a,interferon alfa-2b, interferon alfa-n1, interferon alfa-n3, interferonbeta I a, and interferon gamma-I b; GM-CF and GM-CSF; and EPO.

Particular proteins that can be used in the methods and compositions ofthe invention include, but are not limited to: filgrastim, which is soldin the United States under the trade name Neupogen® (Amgen, ThousandOaks, Calif.); sargramostim, which is sold in the United States underthe trade name Leukine® (Immunex, Seattle, Wash.); and recombinant EPO,which is sold in the United States under the trade name Epogen® (Amgen,Thousand Oaks, Calif.).

Recombinant and mutated forms of GM-CSF can be prepared as described inU.S. Pat. Nos. 5,391,485; 5,393,870; and 5,229,496; all of which areincorporated herein by reference. Recombinant and mutated forms of G-CSFcan be prepared as described in U.S. Pat. Nos. 4,810,643; 4,999,291;5,528,823; and 5,580,755; all of which are incorporated herein byreference.

This invention encompasses the use of native, naturally occurring, andrecombinant proteins. The invention further encompasses mutants andderivatives (e.g., modified forms) of naturally occurring proteins thatexhibit, in vivo, at least some of the pharmacological activity of theproteins upon which they are based. Examples of mutants include, but arenot limited to, proteins that have one or more amino acid residues thatdiffer from the corresponding residues in the naturally occurring formsof the proteins. Also encompassed by the term “mutants” are proteinsthat lack carbohydrate moieties normally present in their naturallyoccurring forms (e.g., nonglycosylated forms). Examples of derivativesinclude, but are not limited to, pegylated derivatives and fusionproteins, such as proteins formed by fusing IgG1 or IgG3 to the proteinor active portion of the protein of interest. See, e.g., Penichet, M. L.and Morrison, S. L., J. Immunol. Methods 248:91-101 (2001).

Antibodies that can be used in combination with compounds of theinvention include monoclonal and polyclonal antibodies. Examples ofantibodies include, but are not limited to, trastuzumab (Herceptin®),rituximab (Rituxan®), bevacizumab (Avastin™), pertuzumab (Omnitarg™),tositumomab (Bexxar°), edrecolomab (Panorex®), and G250. Compounds ofthe invention can also be combined with, or used in combination with,anti-TNF-αantibodies.

Large molecule active agents may be administered in the form ofanti-cancer vaccines. For example, vaccines that secrete, or cause thesecretion of, cytokines such as IL-2, G-CSF, and GM-CSF can be used inthe methods, pharmaceutical compositions, and kits of the invention.See, e.g., Emens, L. A., et al., Curr. Opinion Mol. Ther. 3(1):77-84(2001).

In one embodiment of the invention, the large molecule active agentreduces, eliminates, or prevents an adverse effect associated with theadministration of an immunomodulatory compound of the invention.Depending on the particular immunomodulatory compound of the inventionand the disease or disorder begin treated, adverse effects can include,but are not limited to, drowsiness and somnolence, dizziness andorthostatic hypotension, neutropenia, infections that result fromneutropenia, increased HIV-viral load, bradycardia, Stevens-JohnsonSyndrome and toxic epidermal necrolysis, and seizures (e.g., grand malconvulsions). A specific adverse effect is neutropenia.

Second active agents that are small molecules can also be used toalleviate adverse effects associated with the administration of animmunomodulatory compound of the invention. However, like some largemolecules, many are believed to be capable of providing a synergisticeffect when administered with (e.g., before, after or simultaneously) animmunomodulatory compound of the invention. Examples of small moleculesecond active agents include, but are not limited to anti-cancer agents,antibiotics, immunosuppressive agents, and steroids.

Examples of anti-cancer agents include, but are not limited to acivicinaclarubicin; acodazole hydrochloride; acronine; adozelesin; aldesleukin;altretamine; ambomycin; ametantrone acetate; amsacrine; anastrozole;anthramycin; asparaginase; asperlin; azacitidine; azetepa; azotomycin;batimastat; benzodepa; bicalutamide; bisantrene hydrochloride; bisnafidedimesylate; bizelesin; bleomycin sulfate; brequinar sodium; bropirimine;busulfan; cactinomycin; calusterone; caracemide; carbetimer;carboplatin; carmustine; carubicin hydrochloride; carzelesin;cedefingol; celecoxib (COX-2 inhibitor); chlorambucil; cirolemycin;cisplatin; cladribine; crisnatol mesylate; cyclophosphamide; cytarabine;dacarbazine; dactinomycin; daunorubicin hydrochloride; decitabine;dexormaplatin; dezaguanine; dezaguanine mesylate; diaziquone; docetaxel;doxorubicin; doxorubicin hydrochloride; droloxifene; droloxifenecitrate; dromostanolone propionate; duazomycin; edatrexate; eflornithinehydrochloride; elsamitrucin; enloplatin; enpromate; epipropidine;epirubicin hydrochloride; erbulozole; esorubicin hydrochloride;estramustine; estramustine phosphate sodium; etanidazole; etoposide;etoposide phosphate; etoprine; fadrozole hydrochloride; fazarabine;fenretinide; floxuridine; fludarabine phosphate; fluorouracil;flurocitabine; fosquidione; fostriecin sodium; gemcitabine; gemcitabinehydrochloride; hydroxyurea; idarubicin hydrochloride; ifosfamide;ilmofosine; iproplatin; irinotecan; irinotecan hydrochloride; lanreotideacetate; letrozole; leuprolide acetate; liarozole hydrochloride;lometrexol sodium; lomustine; losoxantrone hydrochloride; masoprocol;maytansine; mechlorethamine hydrochloride; megestrol acetate;melengestrol acetate; melphalan; menogaril; mercaptopurine;methotrexate; methotrexate sodium; metoprine; meturedepa; mitindomide;mitocarcin; mitocromin; mitogillin; mitomalcin; mitomycin; mitosper;mitotane; mitoxantrone hydrochloride; mycophenolic acid; nocodazole;nogalamycin; ormaplatin; oxisuran; paclitaxel; pegaspargase; peliomycin;pentamustine; peplomycin sulfate; perfosfamide; pipobroman; piposulfan;piroxantrone hydrochloride; plicamycin; plomestane; porfimer sodium;porfiromycin; prednimustine; procarbazine hydrochloride; puromycin;puromycin hydrochloride; pyrazofurin; riboprine; safingol; safingolhydrochloride; semustine; simtrazene; sparfosate sodium; sparsomycin;spirogermanium hydrochloride; spiromustine; spiroplatin; streptonigrin;streptozocin; sulofenur; talisomycin; tecogalan sodium; taxotere;tegafur; teloxantrone hydrochloride; temoporfin; teniposide; teroxirone;testolactone; thiamiprine; thioguanine; thiotepa; tiazofurin;tirapazamine; toremifene citrate; trestolone acetate; triciribinephosphate; trimetrexate; trimetrexate glucuronate; triptorelin;tubulozole hydrochloride; uracil mustard; uredepa; vapreotide;verteporfin; vinblastine sulfate; vincristine sulfate; vindesine;vindesine sulfate; vinepidine sulfate; vinglycinate sulfate;vinleurosine sulfate; vinorelbine tartrate; vinrosidine sulfate;vinzolidine sulfate; vorozole; zeniplatin; zinostatin; and zorubicinhydrochloride.

Other anti-cancer drugs include, but are not limited to: 20-epi-1,25dihydroxyvitamin D3; 5-ethynyluracil; abiraterone; aclarubicin;acylfulvene; adecypenol; adozelesin; aldesleukin; ALL-TK antagonists;altretamine; ambamustine; amidox; amifostine; aminolevulinic acid;amrubicin; amsacrine; anagrelide; anastrozole; andrographolide;angiogenesis inhibitors; antagonist D; antagonist G; antarelix;antidorsalizing morphogenetic protein-1; antiandrogen, prostaticcarcinoma; antiestrogen; antineoplaston; antisense oligonucleotides;aphidicolin glycinate; apoptosis gene modulators; apoptosis regulators;apurinic acid; ara-CDP-DL-PTBA; arginine deaminase; asulacrine;atamestane; atrimustine; axinastatin 1; axinastatin 2; axinastatin 3;azasetron; azatoxin; azatyrosin; baccatin III derivatives; balanol;batimastat; BCR/ABL antagonists; benzochlorins; benzoylstaurosporine;beta lactam derivatives; beta-alethine; betaclamycin; B; betulinic acid;bFGF inhibitor; bicalutamide; bisantrene; bisazirdinylspermine;bisnafide; bistratene A; bizelesin; breflate; bropirimine; budotitane;buthionine sulfoximine; calcipotriol; calphostin C; camptothecinderivatives; capecitabine; carboxamide-aminotriazole;carboxyamidotriazole; CaRest M3; CARN 700; cartilage derived inhibitor;carzelesin; casein kinase inhibitors (ICOS); castanospermine; cecropinB; cetrorelix; chlorins; chloroquinoxaline sulfonamide; cicaprost;cis-porphyrin; cladribine; clomifene analogues; clotrimazole;collismycin A; collismycin B; combretastatin A4; combretastatinanalogue; conagenin; crambescidin 816; crisnatol; cryptophycin 8;cryptophycin 8; cryptophycin A derivatives; curacin A;cyclopentanthraquinones; cycloplatam; cypemycin; cytarabine ocfosfate;cytolytic factor; cytostatin; dacliximab; decitabine; dehydrodidemnin B;deslorelin; dexamethasone; dexifosfamide; dexrazoxane; dexverapamil;diaziquone; didemnin B; didox; diethylnorspermine;dihydro-5-azacytidine; dihydrotaxol, 9-; dioxamycin; diphenylspiromustine; docetaxel; docosanol; dolasetron; doxifluridine;doxorubicin; droloxifene; dronabinol; duocarmycin SA; ebselen;ecomustine; edelfosine; edrecolomab; eflornithine; elemene; emitefur;epirubicin; epristeride; estramustine analogue; estrogen agonists;estrogen antagonists; etanidazole; etoposide phosphate; exemestane;fadrozole; fazarabine; fenretinide; filgrastim; finasteride;flavopiridol; flezelastine; fluasterone; fudarabine; fluorodaunorunicinhydrochloride; forfenimex; formestane; fostriecin; fotemustine;gadolinium texaphyrin; gallium nitrate; galocitabine; ganirelix;gelatinase inhibitors; gemcitabine; glutathione inhibitors; hepsulfam;heregulin; hexamethylene bisacetamide; hypericin; ibandronic acid;idarubicin; idoxifene; idramantone; ilmofosine; ilomastat; imatinib(e.g., Gleevec®); imiquimod; immunostimulant peptides; insulin-likegrowth factor-1 receptor inhibitor; interferon agonists; interferons;interleukins; iobenguane; iododoxorubicin; ipomeanol, 4-; iroplact;irsogladine; isobengazole; isohomohalicondrin B; itasetron;jasplakinolide; kahalalide F; lamellarin-N triacetate; lanreotide;leinamycin; lenograstim; lentinan sulfate; leptolstatin; letrozole;leukemia inhibiting factor; leukocyte alpha interferon;leuprolide+estrogen+progesterone; leuprorelin; levamisole; liarozole;linear polyamine analogue; lipophilic disaccharide peptide; lipophilicplatinum compounds; lissoclinamide 7; lobaplatin; lombricin; lometrexol;lonidamine; losoxantrone; loxoribine; lurtotecan; lutetium texaphyrin;lysofylline; lytic peptides; maitansine; mannostatin A; marimastat;masoprocol; maspin; matrilysin inhibitors; matrix metalloproteinaseinhibitors; menogaril; merbarone; meterelin; methioninase;metoclopramide; MIF inhibitor; mifepristone; miltefosine; mirimostim;mitoguazone; mitolactol; mitomycin analogues; mitonafide; mitotoxinfibroblast growth factor-saporin; mitoxantrone; mofarotene;molgramostim;Erbitus, human chorionic gonadotrophin; monophosphoryllipid A+myobacterium cell wall sk; mopidamol; mustard anticancer agent;mycaperoxide B; mycobacterial cell wall extract; myriaporone;N-acetyldinaline; N-substituted benzamides; nafarelin; nagrestip;naloxone+pentazocine; napavin; naphterpin; nartograstim; nedaplatin;nemorubicin; neridronic acid; nilutamide; nisamycin; nitric oxidemodulators; nitroxide antioxidant; nitrullyn; oblimersen (Genasense®);O⁶-benzylguanine; octreotide; okicenone; oligonucleotides; onapristone;ondansetron; ondansetron; oracin; oral cytokine inducer; ormaplatin;osaterone; oxaliplatin; oxaunomycin; paclitaxel; paclitaxel analogues;paclitaxel derivatives; paulauamine; palmitroylrhizoxin; pamidronicacid; panaxytriol; panomifene; parabactin; pazelliptine; pegaspargase;peldesine; pentosan polysulfate sodium; pentostatin; pentrozole;perflubron; perfosfamide; perillyl alcohol; phenazimomycin;phenylacetate; phosphatase inhibitors; picibanil; pilocarpinehydrochloride; pirarubicin; piritrexim; placetin A; placetin B;plasminogen activator inhibitor; platinum complex; platinum compounds;platinum-triamine complex; porfimer sodium; porfiromycin; prednisone;propyl bis-acridone; prostaglandin J2; proteasome inhibitors; proteinA-based immune modulator; protein kinase C inhibitor; protein kinase Cinhibitors, microalgal; protein tyrosine phosphatase inhibitors; purinenucleoside phosphorylase inhibitors; purpurins; pyrazoloacridine;pyridoxylated hemoglobin polyoxyethylene conjugate; raf antagonists;raltitrexed; ramosetron; ras farnesyl protein transferase inhibitors;ras inhibitors; ras-GAP inhibitor; retelliptine demethylated; rhenium Re186 etidronate; rhizoxin; ribozymes; RII retinamide; rohitukine;romurtide; roquinimex; rubiginone B1; ruboxyl; safingol; saintopin;SarCNU; sarcophytol A; sargramostim; Sdi 1 mimetics; semustine;senescence derived inhibitor 1; sense oligonucleotides; signaltransduction inhibitors; sizofiran; sobuzonxane; sodium borocaptate;sodium phenylacetate; solverol; somatomedin binding protein; sonermin;sparfosic acid; spicamycin D; spiromustine; splenopentin; spongistatinI; squalamine; stipiamide; stromelysin inhibitors; sulfinosine;superactive vasoactive intestinal peptide antagonist; suradista;suramin; swainsonine; tallimustine; tamoxifen methiodide; tauromustine;tazarotene; tecogalan sodium; tegafur; tellurapyrylium; telomeraseinhibitors; temoporfin; teniposide; tetrachlorodecaoxide; tetrazomine;thaliblastine; thiocoraline; thrombopoietin; thrombopoietin mimetic;thymalfasin; thymopoietin receptor agonist; thymotrinan; thyroidstimulating hormone; tin ethyl etiopurpurin; tirapazamine; titanocenebichloride; topsentin; toremifene; translation inhibitors; tretinoin;triacetyluridine; triciribine; trimetrexate; triptorelin; tropisetron;turosteride; tyrosine kinase inhibitors; tyrphostins; UBC inhibitors;ubenimex; urogenital sinus-derived growth inhibitory factor; urokinasereceptor antagonists; vapreotide; variolin B; velaresol; veramine;verdins; verteporfin; vinorelbine; vinxaltine; vitaxin; vorozole;zanoterone; zeniplatin; zilascorb; and zinostatin stimalamer.

Specific second active agents include, but are not limited to,rituximab, oblimersen (Genasense®), remicade, docetaxel, celecoxib,melphalan, dexamethasone (Decadron®), steroids, gemcitabine,cisplatinum, temozolomide, etoposide, cyclophosphamide, temodar,carboplatin, procarbazine, gliadel, tamoxifen, topotecan, methotrexate,Arisa®, taxol, taxotere, fluorouracil, leucovorin, irinotecan, xeloda,CPT-11, interferon alpha, pegylated interferon alpha (e.g., PEGINTRON-A), capecitabine, cisplatin, thiotepa, fludarabine, carboplatin,liposomal daunorubicin, cytarabine, doxetaxol, pacilitaxel, vinblastine,IL-2, GM-CSF, dacarbazine, vinorelbine, zoledronic acid, palmitronate,biaxin, busulphan, prednisone, bisphosphonate, arsenic trioxide,vincristine, doxorubicin (Doxil®), paclitaxel, ganciclovir, adriamycin,estramustine sodium phosphate (Emcyt®), sulindac, and etoposide.

5.3 Methods of Treatments and Prevention

Methods of this invention encompass methods of treating, preventing ormanaging various types of cancers. In a preferred embodiment, methods ofthis invention encompass methods of treating, preventing or managingvarious types of leukemias such as chronic lymphocytic leukemia, chronicmyelocytic leukemia, acute lymphoblastic leukemia, acute myelogenousleukemia, and acute myeloblastic leukemia.

As used herein, unless otherwise specified, the term “treating” refersto the administration of a compound of the invention, or otheradditional active agent, after the onset of symptoms of the particulardisease or disorder. As used herein, unless otherwise specified, theterm “preventing” refers to the administration prior to the onset ofsymptoms, particularly to patients at risk of cancer, and in particularleukemia. The term “prevention” includes the inhibition of a symptom ofthe particular disease or disorder. Patients with familial history ofcancer or leukemia in particular are preferred candidates for preventiveregimens. As used here and unless otherwise indicated, the term“managing” encompasses preventing the recurrence of the particulardisease or disorder in a patient who had suffered from it, lengtheningthe time a patient who had suffered from the disease or disorder remainsin remission, and/or reducing mortality rates of the patients.

As used herein, the term “cancer” includes, but is not limited to, solidtumors and blood born tumors. The term “cancer” refers to disease ofskin tissues, organs, blood, and vessels, including, but not limited to,cancers of the bladder, bone or blood, brain, breast, cervix, chest,colon, endrometrium, esophagus, eye, head, kidney, liver, lymph nodes,lung, mouth, neck, ovaries, pancreas, prostate, rectum, stomach, testis,throat, and uterus.

The term “leukemia” refers malignant neoplasms of the blood-formingtissues. The leukemia includes, but is not limited to, chroniclymphocytic leukemia, chronic myelocytic leukemia, acute lymphoblasticleukemia, acute myelogenous leukemia and acute myeloblastic leukemia.The leukemia can be relapsed, refractory or resistant to conventionaltherapy. The term “relapsed” refers to a situation where patients whohave had a remission of leukemia after therapy have a return of leukemiacells in the marrow and a decrease in normal blood cells. The term“refractory or resistant” refers to a circumstance where patients, evenafter intensive treatment, have residual leukemia cells in their narrow.

The various types of the cancers are described in U.S. provisionalapplication No. 60/380,842, filed May 17, 2002, the entireties of whichare incorporated herein by reference (see, e.g., Section 2.2. Types ofCancers). Specific cancer include, but are not limited to, leukemiassuch as chronic lymphocytic leukemia, chronic myelocytic leukemia, acutelymphoblastic leukemia, acute myelogenous leukemia, and acutemyeloblastic leukemia; advanced malignancy, amyloidosis, neuroblastoma,meningioma, hemangiopericytoma, multiple brain metastase, glioblastomamultiforms, glioblastoma, brain stem glioma, poor prognosis malignantbrain tumor, malignant glioma, recurrent malignant giolma, anaplasticastrocytoma, anaplastic oligodendroglioma, neuroendocrine tumor, rectaladenocarcinoma, Dukes C & D colorectal cancer, unresectable colorectalcarcinoma, metastatic hepatocellular carcinoma, Kaposi's sarcoma,karotype acute myeloblastic leukemia, Hodgkin's lymphoma, non-Hodgkin'slymphoma, cutaneous T-Cell lymphoma, cutaneous B-Cell lymphoma, diffuselarge B-Cell lymphoma, low grade follicular lymphoma, malignantmelanoma, malignant mesothelioma, malignant pleural effusionmesothelioma syndrome, peritoneal carcinoma, papillary serous carcinoma,gynecologic sarcoma, soft tissue sarcoma, scleroderma, cutaneousvasculitis, Langerhans cell histiocytosis, leiomyosarcoma,fibrodysplasia ossificans progressive, hormone refractory prostatecancer, resected high-risk soft tissue sarcoma, unrescectablehepatocellular carcinoma, Waldenstrom's macroglobulinemia, smolderingmyeloma, indolent myeloma, fallopian tube cancer, androgen independentprostate cancer, androgen dependent stage IV non-metastatic prostatecancer, hormone-insensitive prostate cancer, chemotherapy-insensitiveprostate cancer, papillary thyroid carcinoma, follicular thyroidcarcinoma, medullary thyroid carcinoma, and lciomyoma. In oneembodiment, the cancer is primary or metastatic. In another embodiment,the cancer is relapsed, refractory or resistance to chemotherapy orradiation; in particular, refractory to thalidomide. As used herein, theterm “cancer” does not include myelodysplastic syndromes or MDS.

This invention encompasses methods of treating patients who have beenpreviously treated for cancer, but are non-responsive to standardtherapies, as well as those who have not previously been treated. Theinvention also encompasses methods of treating patients regardless ofpatient's age, although some diseases or disorders are more common incertain age groups. The invention further encompasses methods oftreating patients who have undergone surgery in an attempt to treat thedisease or condition at issue, as well as those who have not. Becausepatients with cancer have heterogenous clinical manifestations andvarying clinical outcomes, the treatment given to a patient may vary,depending on his/her prognosis. The skilled clinician will be able toreadily determine without undue experimentation specific secondaryagents, types of surgery, and types of non-drug based standard therapythat can be effectively used to treat an individual patient with cancer.

Methods encompassed by this invention comprise administering one or moreimmunomodulatory compound of the invention, or a pharmaceuticallyacceptable salt, solvate, hydrate, stereoisomer, clathrate, or prodrugthereof, to a patient (e.g., a human) suffering, or likely to suffer,from cancer, particularly leukemia.

In one embodiment of the invention, an immunomodulatory compound of theinvention can be administered orally and in single or divided dailydoses in an amount of from about 0.10 to about 150 mg/day. In apreferred embodiment,3-(4-amino-1-oxo-1,3-dihydro-isoindol-2-yl-piperidine-2,6-dione(Revlimid®) may be administered in an amount of from about 0.10 to 150mg per day, from about 1 to about 50 mg per day, or from about 5 toabout 25 mg per day. Specific doses per day include 1, 2, 3, 4, 5, 6, 7,8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25,26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43,44, 45, 46, 47, 48, 49 or 50 mg per day.

In a preferred embodiment3-(4-amino-1-oxo-1,3-dihydro-isoindol-2-yl-piperidine-2,6-dione(Revlimid®) may be administered in an amount of from about 1 to 50 mgper day, or from about 5 to about 25 mg per day to patients with varioustypes of leukemias such as chronic lymphocytic leukemia, and acutemyeloblastic leukemia.

In particular,3-(4-amino-1-oxo-1,3-dihydro-isoindol-2-yl-piperidine-2,6-dione(Revlimid®) may be administered to patients with chronic lymphocyticleukemia in an amount of from about 1 to 50 mg per day, or from about 5to about 25 mg per day. In an specific embodiment,3-(4-amino-1-oxo-1,3-dihydro-isoindol-2-yl-piperidine-2,6-dione(Revlimid®) may be administered to patients with chronic lymphocyticleukemia in an amount of about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12,13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30,31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48,49 or 50 mg per day. In a specific embodiment, Revlimid® can beadministered in an amount of about 25 mg/day to patients with chroniclymphocytic leukemia.

In one embodiment, the recommended starting dose of3-(4-amino-1-oxo-1,3-dihydro-isoindol-2-yl-piperidine-2,6-dione(Revlimid®) is 10 mg per day. The dose can be escalated every week to15, 20, 25, 30, 35, 40, 45 and 50 mg/day. The patients who are dosedinitially to 10 mg and who experience thrombocytopenia or neutropeniathat develops within or after the first four weeks of starting Revlimid®therapy may have their dosage adjusted according to a platelet count orabsolute neutrophil count (ANC).

-   -   5.31 Combination Therapy with a Second Active Agent

Specific methods of the invention comprise administering animmunomodulatory compound of the invention, or a pharmaceuticallyacceptable salt, solvate, hydrate, steroisomer, clathrate, or prodrugthereof, in combination with one or more second active agents, and/or incombination with radiation therapy, blood transfusions, or surgery.Examples of immunomodulatory compounds of the invention are disclosedherein (see, e.g., section 5.1). Examples of second active agents arealso disclosed herein (see, e.g., section 5.2).

Administration of an immunomodulatory compound of the invention and thesecond active agents to a patient can occur simultaneously orsequentially by the same or different routes of administration. Thesuitability of a particular route of administration employed for aparticular active agent will depend on the active agent itself (e.g.,whether it can be administered orally without decomposing prior toentering the blood stream) and the disease being treated. A preferredroute of administration for an immunomodulatory compound of theinvention is orally. Preferred routes of administration for the secondactive agents or ingredients of the invention are known to those ofordinary skill in the art. See, e.g., Physicians' Desk Reference1755-1760 (56^(th) ed., 2002).

In one embodiment of the invention, the second active agent isadministered intravenously or subcutaneously and once or twice daily inan amount of from about 1 to about 1,000 mg, from about 5 to about 500mg, from about 10 to about 375 mg, or from about 50 to about 200 mg. Thespecific amount of the second active agent will depend on the specificagent used, the type of disease being treated or managed, the severityand stage of disease, and the amount(s) of immunomodulatory compounds ofthe invention and any optional additional active agents concurrentlyadministered to the patient. In a particular embodiment, the secondactive agent is rituximab, oblimersen (Genasense®), GM-CSF, G-CSF, EPO,taxotere, irinotecan, dacarbazine, transretinoic acid, topotecan,pentoxifylline, ciprofloxacin, dexamethasone, vincristine, doxorubicin,COX-2 inhibitor, IL2, IL8, IL18, IFN, Ara-C, vinorelbine, or acombination thereof.

In a specific embodiment, an immunomodulatory compound of the inventionis administered in combination with rituximab to patients withleukemias. In a specific embodiment, Revlimid® is administered in anamount of from about 5 to about 25 mg per day to patients with chroniclymphocytic leukemia in combination with rituximab in an amount of 375mg/m².

In another embodiment, an immunomodulatory compound of the invention isadministered in combination with fludarabine, carboplatin, and/ortopetecan to patients with refractory or relapsed or high-risk acutemyelogenous leukemia.

In another embodiment, an immunomodulatory compound of the invention isadministered in combination with liposomal daunorubicin, topotecanand/or cytarabine to patients with unfavorable karotype acutemyeloblastic leukemia.

In another embodiment, an immunomodulatory compound of the invention isadministered alone or in combination with a second active ingredientsuch as vinblastine or fludarabine to patients with various types oflymphoma, including, but not limited to, Hodgkin's lymphoma,non-Hodgkin's lymphoma, cutaneous T-Cell lymphoma, cutaneous B-Celllymphoma, diffuse large B-Cell lymphoma or relapsed or refractory lowgrade follicular lymphoma.

In another embodiment, GM-CSF, G-CSF or EPO is administeredsubcutaneously during about five days in a four to six week cycle in anamount of from about 1 to about 750 mg/m²/day, preferably in an amountof from about 25 to about 500 mg/m²/day, more preferably in an amount offrom about 50 to about 250 mg/m²/day, and most preferably in an amountof from about 50 to about 200 mg/m²/day. In a certain embodiment, GM-CSFmay be administered in an amount of from about 60 to about 500 mcg/m²intravenously over 2 hours, or from about 5 to about 12 mcg/m²/daysubcutaneously. In a specific embodiment, G-CSF may be administeredsubcutaneously in an amount of about 1 mcg/kg/day initially and can beadjusted depending on rise of total granulocyte counts. The maintenancedose of G-CSF may be administered in an amount of about 300 (in smallerpatients) or 480 mcg subcutaneously. In a certain embodiment, EPO may beadministered subcutaneously in an amount of 10,000 Unit 3 times perweek.

This invention also encompasses a method of increasing the dosage of ananti-cancer drug or agent that can be safely and effectivelyadministered to a patient, which comprises administering to a patient(e.g., a human) an immunomodulatory compound of the invention, or apharmaceutically acceptable derivative, salt, solvate, clathrate,hydrate, or prodrug thereof. Patients that can benefit by this methodare those likely to suffer from an adverse effect associated withanti-cancer drugs for treating a specific cancer of the blood, skin,subcutaneous tissue, lymph nodes, brain, lung, liver, bone, intestine,colon, heart, pancreas, adrenal, kidney, prostate, breast colorectal, orcombinations thereof. The administration of an immunomodulatory compoundof the invention alleviates or reduces adverse effects which are of suchseverity that it would otherwise limit the amount of anti-cancer drug.

In one embodiment, an immunomodulatory compound of the invention can beadministered orally and daily in an amount of from about 0.10 to about150 mg, and preferably from about 1 to about 50 mg, more preferably fromabout 5 to about 25 mg prior to, during, or after the occurrence of theadverse effect associated with the administration of an anti-cancer drugto a patient. In a particular embodiment, an immunomodulatory compoundof the invention is administered in combination with specific agentssuch as heparin, aspirin, coumadin, or G-CSF to avoid adverse effectsthat are associated with anti-cancer drugs such as but not limited toneutropenia or thrombocytopenia.

In another embodiment, this invention encompasses a method of treating,preventing and/or managing cancer, which comprises administering animmunomodulatory compound of the invention, or a pharmaceuticallyacceptable salt, solvate, hydrate, stereoisomer, clathrate, or prodrugthereof, in conjunction with (e.g., before, during, or after)conventional therapy including, but not limited to, surgery,immunotherapy, biological therapy, radiation therapy, or other non-drugbased therapy presently used to treat, prevent or manager cancer. Thecombined use of the immunomodulatory compounds of the invention andconventional therapy may provide a unique treatment regimen that isunexpectedly effective in certain patients. Without being limited bytheory, it is believed that immunomodulatory compounds of the inventionmay provide additive or synergistic effects when given concurrently withconventional therapy.

As discussed elsewhere herein, the invention encompasses a method ofreducing, treating and/or preventing adverse or undesired effectsassociated with conventional therapy including, but not limited to,surgery, chemotherapy, radiation therapy, hormonal therapy, biologicaltherapy and immunotherapy. An immunomodulatory compound of the inventionand other active ingredient can be administered to a patient prior to,during, or after the occurrence of the adverse effect associated withconventional therapy.

In one embodiment, an immunomodulatory compound of the invention can beadministered in an amount of from about 0.10 to about 150 mg, andpreferably from about 1 to about 50 mg, more preferably from about 5 toabout 25 mg orally and daily alone, or in combination with a secondactive agent disclosed here (see, e.g., section 5.2), prior to, during,or after the use of conventional therapy.

-   -   5.3.2 Use with Transplantation Therapy

Compounds of the invention can be used to reduce the risk of GraftVersus Host Disease (GVHD). Therefore, the invention encompasses amethod of treating, preventing and/or managing cancer, which comprisesadministering the immunomodulatory compound of the invention, or apharmaceutically acceptable salt, solvate, hydrate, stereoisomer,clathrate, or prodrug thereof, in conjunction with transplantationtherapy.

As those of ordinary skill in the art are aware, the treatment of canceris often based on the stages and mechanism of the disease. For example,as inevitable leukemic transformation develops in certain stages ofcancer, transplantation of peripheral blood stem cells, hematopoieticstem cell preparation or bone marrow may be necessary. The combined useof the immunomodulatory compound of the invention and transplantationtherapy provides a unique and unexpected synergism. In particular, animmunomodulatory compound of the invention exhibits immunomodulatoryactivity that may provide additive or synergistic effects when givenconcurrently with transplantation therapy in patients with cancer.

An immunomodulatory compound of the invention can work in combinationwith transplantation therapy reducing complications associated with theinvasive procedure of transplantation and risk of GVHD. This inventionencompasses a method of treating, preventing and/or managing cancerwhich comprises administering to a patient (e.g., a human) animmunomodulatory compound of the invention, or a pharmaceuticallyacceptable salt, solvate, hydrate, stereoisomer, clathrate, or prodrugthereof, before, during, or after the transplantation of umbilical cordblood, placental blood, peripheral blood stem cell, hematopoietic stemcell preparation or bone marrow. Examples of stem cells suitable for usein the methods of the invention are disclosed in U.S. patent publicationnos. 2002/0123141, 2003/0235909 and 2003/0032179, by R. Hariri et al.,the entireties of which are incorporated herein by reference.

In one embodiment of this method, and immunomodulatory compound of theinvention is administered to patients with leukemias before, during, orafter the transplantation of autologous peripheral blood progenitorcell.

In another embodiment, an immunomodulatory compound of the invention isadministered to patients with relapsed leukemia after the stem celltransplantation.

-   -   5.3.3 Cycling Therapy

In certain embodiments, the prophylactic or therapeutic agents of theinvention are cyclically administered to a patient. Cycling therapyinvolves the administration of an active agent for a period of time,followed by a rest for a period of time and repeating this sequentialadministration. Cycling therapy can reduce the development of resistanceto one or more of the therapies, avoid or reduce the side effects of oneof the therapies, and/or improves the efficacy of the treatment.

Consequently, in one specific embodiment of the invention, animmunomodulatory compound of the invention is administered daily in asingle or divided doses in a four to six week cycle with a rest periodof about a week or two weeks. The invention further allows thefrequency, number, and length of dosing cycles to be increased. Thus,another specific embodiment of the invention encompasses theadministration of an immunomodulatory compound of the invention for morecycles than are typical when it is administered alone. In yet anotherspecific embodiment of the invention, an immunomodulatory compound ofthe invention is administered for a greater number of cycles that wouldtypically cause dose-limiting toxicity in a patient to whom a secondactive ingredient is not also being administered.

In one embodiment, an immunomodulatory compound of the invention isadministered daily and continuously for three or four weeks at a dose offrom about 0.10 to about 150 mg/d followed by a break of one or twoweeks. In a particular embodiment, an immunomodulatory compound of theinvention is administered in an amount of from about 1 to about 50mg/day, preferably in an amount of about 25 mg/day for three to fourweeks, followed by one week or two weeks of rest in a four or six weekcycle.

In a preferred embodiment,3-(4-amino-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione(Revlimid®) is administered to patients with leukemia in an amount offrom about 0.10 to about 150 mg per day for 21 days followed by sevendays rest in a 28 day cycle. In the most preferred embodiment,3-(4-amino-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione(Revlimid®) is administered to patients with refractory or relapsedchronic lymphocytic leukemia in an amount of about 25 mg per day for 21days followed by seven days rest in a 28 day cycle.

In one embodiment of the invention, an immunomodulatory compound of theinvention and a second active ingredient are administered orally, withadministration of an immunomodulatory compound of the inventionoccurring 30 to 60 minutes prior to a second active ingredient, during acycle of four to six weeks. In another embodiment of the invention, animmunomodulatory compound of the invention is administered orally and asecond active ingredient is administered by intravenous infusion.

In a specific embodiment, one cycle comprises the administration of fromabout 10 to about 25 mg/day of Revlimid® and from about 50 to about 750mg/m²/day of a second active ingredient daily for three to four weeksand then one or two weeks of rest. In a preferred embodiment, rituximabcan be administered in an amount of 375 mg/m² as an additional activeagent to patients with refractory or relapsed chronic lymphocyticleukemia. Typically, the number of cycles during which the combinatorialtreatment is administered to a patient will be from about one to about24 cycles, more typically from about two to about 16 cycles, and evenmore typically from about four to about three cycles.

5.4 Pharmaceutical Compositions and Dosage Forms

Pharmaceutical compositions can be used in the preparation ofindividual, single unit dosage forms. Pharmaceutical compositions anddosage forms of the invention comprise an immunomodulatory compound ofthe invention, or a pharmaceutically acceptable salt, solvate, hydrate,stereisomer, clathrate, or prodrug thereof. Pharmaceutical compositionsand dosage forms of the invention can further comprise one or moreexcipients.

Pharmaceutical compositions and dosage forms of the invention can alsocomprise one or more additional active ingredients. Consequently,pharmaceutical compositions and dosage forms of the invention comprisethe active ingredients disclosed herein (e.g., and immunomodulatorycompound of the invention and a second active agent). Examples ofoptional second, or additional, active ingredients are disclosed herein(see, e.g., section 5.2).

Single unit dosage forms of the invention are suitable for oral, mucosal(e.g., nasal, sublingual, vaginal, buccal, or rectal), parenteral (e.g.,subcutaneous, intravenous, bolus injection, intramuscular, orintraarterial), topical (e.g., eye drops or other opthalmicpreparations), transdermal or transcutaneous administration to apatient. Examples of dosage forms include, but are not limited to:tablets, caplets; capsules, such as soft elastic gelatin capsules;cachets; troches; lozenges; dispersions; suppositories; powders;aerosols (e.g., nasal sprays or inhalers); gels; liquid dosage formssuitable for oral or mucosal administration to a patient, includingsuspensions (e.g., aqueous or non-aqueous liquid suspensions,oil-in-water emulsions, or a water-in-oil liquid emulsions), solutions,and elixirs; liquid dosage forms suitable for parenteral administrationto a patient; eye drops or other opthalmic preparations suitable fortopical administration; and sterile solids (e.g., crystalline oramorphous solids) that can be reconstituted to provide liquid dosageforms suitable for parenteral administration to a patient.

The composition, shape, and type of dosage forms of the invention willtypically vary depending on their use. For example, a dosage form usedin the acute treatment of a disease may contain larger amounts of one ormore of the active ingredients it comprises than a dosage form used inthe chronic treatment of the same disease. Similarly, a parenteraldosage form may contain smaller amounts of one or more of the activeingredients it comprises than an oral dosage form used to treat the samedisease. These and other ways in which specific dosage forms encompassedby this invention will vary from one another will be readily apparent tothose skilled in the art. See., e.g., Remington's PharmaceuticalSciences, 18th ed., Mack Publishing, Easton, Pa. (1990).

Typical pharmaceutical compositions and dosage forms comprise one ormore excipients. Suitable excipients are well know to those skilled inthe art of pharmacy, and non-limiting examples of suitable excipientsare provided herein. Whether a particular excipient is suitable forincorporation into a pharmaceutical composition or dosage form dependson a variety of factors well known in the art including, but not limitedto, the way in which the dosage form will be administered to a patient.For example, oral dosage forms such as tablets may contain excipientsnot suited for use in parenteral dosage forms. The suitability of aparticular excipient may also depend on the specific active ingredientsin the dosage form. For example, the decomposition of some activeingredients may be accelerated by some excipients such as lactose, orwhen exposed to water. Active ingredients that comprise primary orsecondary amines are particularly susceptible to such accelerateddecomposition. Consequently, this invention encompasses pharmaceuticalcompositions and dosage forms that contain little, if any, lactose othermono- or disaccharides. As used herein, the term “lactose-free” meansthat the amount of lactose present, if any, is insufficient tosubstantially increase the degradation rate of an active ingredient.

Lactose-free compositions of the invention can comprise excipients thatare well know in the art and are listed, for example, in the U.S.Pharmacopeia (USP) 25-NF20 (2002). In general, lactose-free compositionscomprise active ingredients, a binder/filler, and a lubricant inpharmaceutically compatible and pharmaceutically acceptable amounts.Preferred lactose-free dosage forms comprise active ingredients,microcrystalline cellulose, pre-gelatinized starch, and magnesiumstearate.

This invention further encompasses anhydrous pharmaceutical compositionsand dosage forms comprising active ingredients, since water canfacilitate the degradation of some compounds. For example, the additionof water (e.g., 5%) is widely accepted in the pharmaceutical arts as ameans of simulating long-term storage in order to determinecharacteristics such as shelf-life or the stability of formulations overtime. See, e.g., Jens T. Carstensen, Drug Stability; Principles &Practice, 2d. Ed., Marcel Dekker, NY, N.Y., 1995 pp. 379-80. In effect,water and heat accelerate the decomposition of some compounds. Thus, theeffect of water on a formulation can be of great significance sincemoisture and/or humidity are commonly encountered during manufacture,handling, packaging, storage, shipment, and use of formulations.

Anhydrous pharmaceutical compositions and dosage forms of the inventioncan be prepared using anhydrous or low moisture containing ingredientsand low moisture or low humidity conditions. Pharmaceutical compositionsand dosage forms that comprise lactose and at least one activeingredient that comprises a primary or secondary amine are preferablyanhydrous if substantial contact with moisture and/or humidity duringmanufacturing, packaging, and/or storage is expected.

An anhydrous pharmaceutical composition should be prepared and storedsuch that its anhydrous nature is maintained. Accordingly, anhydrouscompositions are preferably packaged using materials known to preventexposure to water such that they can be included in suitable formularykits. Examples of suitable packaging include, but are not limited to,hermetically sealed foils, plastics, unit dose containers (e.g., vials),blister packs, and strip packs.

The invention further encompasses pharmaceutical compositions and dosageforms that comprise one or more compounds that reduce the rate by whichan active ingredient will decompose. Such compounds, which are referredto herein as “stabilizers,” include, but are not limited to,antioxidants such as ascorbic acid, pH buffers, or salt buffers.

Like the amounts and types of excipients, the amounts and specific typesof active ingredients in a dosage form may differ depending on factorssuch as, but not limited to, the route by which it is to be administeredto patients. However, typical dosage forms of the invention comprise animmunomodulatory compound of the invention or a pharmaceuticallyacceptable salt, solvate, hydrate, stereoisomer, clathrate, or prodrugthereof in an amount of from about 0.10 to about 150 mg. Typical dosageforms comprise an immunomodulatory compound of the invention or apharmaceutically acceptable salt, solvate, hydrate, stereoisomer,clathrate, or prodrug thereof in an amount of about 0.1, 1, 2.5, 5, 7.5,10, 12.5, 15, 17.5, 20, 25, 50, 100, 150 or 200 mg. In a specificembodiment, a preferred dosage form comprises3-(4-amino-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione(Revlimid®) in an amount of about 1, 2.5, 5, 10, 15, 20, 25, or 50 mg.Typical dosage forms comprise the second active ingredient in an amountof 1 to about 1000 mg, from about 5 to about 500 mg, from about 10 toabout 350 mg, or from about 50 to about 200 mg. Of course, the specificamount of the anti-cancer drug will depend on the specific agent used,the type of cancer being treated or managed, and the amount(s) of animmunomodulatory compound of the invention and any optional additionalactive agents concurrently administered to the patient.

-   -   5.4.1 Oral Dosage Forms

Pharmaceutical compositions of the invention that are suitable for oraladministration can be presented as discrete dosage forms, such as, butare not limited to, tablets (e.g., chewable tablets), caplets, capsules,and liquids (e.g., flavored syrups). Such dosage forms containpredetermined amounts of active ingredients, and may be prepared bymethods of pharmacy well known to those skilled in the art. Seegenerally, Remington's Pharmaceutical Sciences, 18th ed., MackPublishing, Easton Pa. (1990).

In one embodiment, a preferred dosage form is a capsule or tablecomprising3-(4-amino-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione(Revlimid®) in an amount of about 1, 2.5, 5, 10, 15, 20, 25 or 50 mg. Ina specific embodiment, a preferred capsule or tablet dosage formcomprises 3-(4-amino-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione(Revlimid®) in an amount of about 5 or 10 mg.

Typical oral dosage forms of the invention are prepared by combining theactive ingredients in an intimate admixture with at least one excipientaccording to conventional pharmaceutical compounding techniques.Excipients can take a wide variety of forms depending on the form ofpreparation desired for administration. For example, excipients suitablefor use in oral liquid or aerosol dosage forms (e.g., powders, tablets,capsules, and caplets) include, but are not limited to, starches,sugars, micro-crystalline cellulose, diluents, granulating agents,lubricants, binders, and disintegrating agents.

Because of their ease of administration, tablets and capsules representthe most advantageous oral dosage unit forms in which case solidexcipients are employed. If desired, tablets can be coated by standardaqueous or nonaqueous techniques. Such dosage forms can be prepared byany of the methods of pharmacy. In general, pharmaceutical compositionsand dosage forms are prepared by uniformly and intimately admixing theactive ingredients with liquid carriers, finely divided solid carriers,or both, and then shaping the product into the desired presentation ifnecessary.

For example, a tablet can be prepared by compression or molding.Compressed tablets can be prepared by compressing in a suitable machinethe active ingredients in a free-flowing form such as powder orgranules, optionally mixed with an excipient. Molded tablets can be madeby molding in a suitable machine a mixture of the powdered compoundmoistened with an inert liquid diluent.

Examples of excipients that can be used in oral dosage forms of theinvention include, but are not limited to, binders, filler,disintegrants, and lubricants. Binders suitable for use inpharmaceutical compositions and dosage forms include, but are notlimited to, corn starch, potato starch, or other starches, gelatin,natural and synthetic gums such as acacia, sodium alginate, alginicacid, other alginates, powdered tragacanth, guar gum, cellulose and itsderivatives (e.g., ethyl cellulose, cellulose acetate, carboxymethylcellulose calcium, sodium carboxymethyl cellulose), polyvinylpyrrolidone, methyl cellulose, pre-gelatinized starch, hydroxypropylmethyl cellulose, (e.g., Nos. 2208, 2906, 2910), microcrystallinecellulose, and mixtures thereof.

Suitable forms of microcrystalline cellulose include, but are notlimited to, the materials sols as AVICEL-PH-101, AVICEL-PH-103 AVICELRC-581, AVICEL-PH-105 (available from FMC Corporation, American ViscoseDivision, Avicel Sales, Marcus Hook, Pa.), and mixtures thereof. Anspecific binder is a mixture of microcrystalline cellulose and sodiumcarboxymethyl cellulose sold as AVICEL RC-581. Suitable anhydrous or lowmoisture excipients or additives include AVICEL-PH-103™ and Starch 1500LM.

Examples of fillers suitable for use in the pharmaceutical compositionsand dosage forms disclosed herein include, but are not limited to, talc,calcium carbonate (e.g., granules or powder), microcrystallinecellulose, powdered cellulose, dextrates, kaolin, mannitol, silicicacid, sorbitol, starch, pre-gelatinized starch, and mixtures thereof.The binder or filler in pharmaceutical compositions of the invention istypically present in from about 50 to about 99 weight percent of thepharmaceutical composition or dosage form.

Disintegrants are used in the compositions of the invention to providetablets that disintegrate when exposed to an aqueous environment.Tablets that contain too much disintegrant may disintegrate in storage,while those that contain too little may not disintegrate at a desiredrate or under the desired conditions. Thus, a sufficient amount ofdisintegrant that is neither too much nor too little to detrimentallyalter the release of the active ingredients should be used to form solidoral dosage forms of the invention. The amount of disintegrant usedvaries based upon the type of formulation, and is readily discernible tothose of ordinary skill in the art. Typical pharmaceutical compositionscomprise from about 0.5 to about 15 weight percent of disintegrant,preferably from about 1 to about 5 weight percent of disintegrant.

Disintegrants that can be used in pharmaceutical compositions and dosageforms of the invention include, but are not limited to, agar-agar,alginic acid, calcium carbonate, microcrystalline cellulose,croscarmellose sodium, crospovidone, polacrilin potassium, sodium starchglycolate, potato or tapioca starch, other starches, pre-gelatinizedstarch, other starches, clays, other algins, other celluloses, gums, andmixtures thereof.

Lubricants that can be used in pharmaceutical compositions and dosageforms of the invention include, but are not limited to, calciumstearate, magnesium stearate, mineral oil, light mineral oil, glycerin,sorbitol, mannitol, polyethylene glycol, other glycols, stearic acid,sodium lauryl sulfate, talc, hydrogenated vegetable oil (e.g., peanutoil, cottonseed oil, sunflower oil, sesame oil, olive oil, corn oil, andsoybean oil), zinc stearate, ethyl oleate, ethyl laureate, agar, andmixtures thereof. Additional lubricants include, for example, a syloidsilica gel (AEROSIL200, manufactured by W.R. Grace Co. of Baltimore,Md.), a coagulated aerosol of synthetic silica (marketed by Degussa Co.of Plano, Tex.), CAB-O-SIL (a pyrogenic silicon dioxide product sold byCabot Co. of Boston, Mass.), and mixtures thereof. If used at alllubricants are typically used in an amount of less than about 1 weightpercent of the pharmaceutical compositions or dosage forms into whichthey are incorporated.

A preferred solid oral dosage form of the invention comprises animmunomodulatory compound of the invention, anhydrous lactose,microcrystalline cellulose, polyvinylpyrrolidone, stearic acid,colloidal anhydrous silica, and gelatin.

-   -   5.4.2 Delayed Release Dosage Forms

Active ingredients of the invention can be administered by controlledrelease means or by delivery devices that are well known to those ofordinary skill in the art. Examples include, but are not limited to,those described in U.S. Pat. Nos. 3,845,770, 3,916,899; 3,536,809;3,598,123; and 4,008,719, 5,674,533, 3,059,595, 5,591,767, 5,120,548,5,073,543, 5,639,476, 5,354,556, and 5,733,566, each of which isincorporated herein by reference. Such dosage forms can be used toprovide slow or controlled-release of one or more active ingredientsusing, for example, hydropropylmethyl cellulose, other polymer matrices,gels, permeable membranes, osmotic systems, multilayer coatings,microparticles, liposomes, microspheres, or a combination thereof toprovide the desired release profile in varying portions. Suitablecontrolled-release formulations known to those of ordinary skill in theart, including those described herein, can be readily selected for usewith the active ingredients of the invention. The invention thusencompasses single unit dosage forms suitable for oral administrationsuch as, but not limited to, tablets, capsules, gelcaps, and capletsthat are adapted for controlled-release.

All controlled-release pharmaceutical products have a common goal ofimproving drug therapy over that achieved by their non-controlledcounterparts. Ideally, the use of an optimally designedcontrolled-release preparation in medical treatment is characterized bya minimum of drug substance being employed to cure or control thecondition in a minimum amount of time. Advantages of controlled-releaseformulations include extended activity of the drug, reduced dosagefrequency, and increased patient compliance. In addition,controlled-release formulations can be used to affect the time of onsetof action or other characteristics, such as blood levels of the drug,and can thus affect the occurrence of side (e.g., adverse) effects.

Most controlled-release formulations are designed to initially releasean amount of drug (active ingredient) that promptly produces the desiredtherapeutic effect, and gradually and continually release of otheramounts of drug to maintain this level of therapeutic or prophylacticeffect over an extended period of time. In order to maintain thisconstant level of drug in the body, the drug must be released from thedosage form at a rate that will replace the amount of drug beingmetabolized and excreted from the body. Controlled-release of an activeingredient can be stimulated by various conditions including, but notlimited to, pH, temperature, enzymes, water, or other physiologicalconditions or compounds.

-   -   5.4.3 Parenteral Dosage Forms

Parenteral dosage forms can be administered to patients by variousroutes including, but not limited to, subcutaneous, intravenous(including bolus injection), intramuscular, and intraarterial. Becausetheir administration typically bypasses patients' natural defensesagainst contaminants, parenteral dosage forms are preferably sterile orcapable of being sterilized prior to administration to a patient.Examples of parenteral dosage forms include, but are not limited to,solutions ready for injection, dry products ready to be dissolved orsuspended in a pharmaceutically acceptable vehicle for injection,suspensions ready for injection, and emulsions.

Suitable vehicles that can be used to provide parenteral dosage forms ofthe invention are well know to those skilled in the art. Examplesinclude, but are not limited to: Water for Injection USP; aqueousvehicles such as, but not limited to, Sodium Chloride Injection,Ringer's Injection, Dextrose Injection, Dextrose and Sodium ChlorideInjection, and Lactated Ringer's Injection; water-miscible vehicles suchas, but not limited to, ethyl alcohol, polyethylene glycol, andpolypropylene glycol; and non-aqueous vehicles such as, but not limitedto, corn oil, cottonseed oil, peanut oil, sesame oil, ethyl olcate,isopropyl myristate, and benzyl benzoate.

Compounds that increase the solubility of one or more of the activeingredients disclosed herein can also be incorporated into theparenteral dosage forms of the invention. For example, cyclodextrin andits derivatives can be used to increase the solubility of animmunomodulatory compound of the invention and its derivatives. See,e.g., U.S. Pat. No. 5,134,127, which is incorporated herein byreference.

-   -   5.4.4 Topical and Mucosal Dosage Forms

Topical and mucosal dosage forms of the invention include, but are notlimited to, sprays, aerosols, solutions, emulsions, suspensions, eyedrops or other ophthalmic preparations, or other forms known to one ofskill in the art. See, e.g., Remington's Pharmaceutical Sciences,16^(th) and 18^(th) eds., Mack Publishing, Easton Pa. (1980 & 1990); andIntroduction to Pharmaceutical Dosage Forms, 4th ed., Lea & Febiger,Philadelphia (1985). Dosage forms suitable for treating mucosal tissueswithin the oral cavity can be formulated as mouthwashes or as oral gels.

Suitable excipients (e.g., carriers and diluents) and other materialsthat can be used to provide topical and mucosal dosage forms encompassedby this invention are well known to those skilled in the pharmaceuticalarts, and depend on the particular tissue to which a givenpharmaceutical composition or dosage form will be applied. With thatfact in mind, typical excipients include, but are not limited to, water,acetone, ethanol, ethylene glycol, propylene glycol, butane-1,3-diol,isopropyl myristate, isopropyl palmitate, mineral oil, and mixturesthereof to form solutions, emulsions or gels, which are non-toxic andpharmaceutically acceptable. Moisturizers or humectants can also beadded to pharmaceutical compositions and dosage forms if desired.Examples of such additional ingredients are well know in the art. See,e.g., Remington's Pharmaceutical Sciences, 16^(th) and 18^(th) eds.,Mack Publishing, Easton Pa. (1980 & 1990).

The pH of a pharmaceutical composition or dosage form may also beadjusted to improve delivery of one or more active ingredients.Similarly, the polarity of a solvent carrier, its ionic strength, ortonicity can be adjusted to improve delivery. Compounds such asstearates can also be added to pharmaceutical compositions or dosageforms to advantageously alter the hydrophilicity or lipophilicity of oneor more active ingredients so as to improve delivery. In this regard,stearates can serve as a lipid vehicle for the formulation, as anemulsifying agent or surfactant, and as a delivery-enhancing orpenetration-enhancing agent. Different salts, hydrates or solvates ofthe active ingredients can be used to further adjust the properties ofthe resulting composition.

6. EXAMPLES

Certain embodiments of the invention are illustrated by the followingnon-limiting example.

6.1 Toxicology Studies

The effects of3-(4-amino-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione(Revlimid®) on cardiovascular and respiratory function are investigatedin anesthetized dogs. Two groups of Beagle dogs (2/sex/group) are used.One group receives three doses of vehicle only and the other receivesthree ascending doses of3-(4-amino-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione (2, 10,and 20 mg/kg). In all cases, doses of3-(4-amino-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione orvehicle are successively administered via infusion through the jugularvein separated by intervals of at least 30 minutes.

The cardiovascular and respiratory changes induced by3-(4-amino-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione areminimal at all doses when compared to the vehicle control group. Theonly statistically significant difference between the vehicle andtreatment groups is a small increase in arterial blood pressure (from 94mmHg to 101 mmHg) following administration of the low dose of3-(4-amino-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione. Thiseffect lasts approximately 15 minutes and is not seen at higher doses.Deviations in femoral blood flow, respiratory parameters, and Qtcinterval are common to both the control and treated groups and are notconsidered treatment-related.

6.2 Clinical Studies in Patients

-   -   6.2.1 Treatment of Chronic Lymphocytic Leukemia

3-(4-amino-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione(Revlimid®) was orally administered to patients with refractory orrelapsed chronic lymphocytic leukemia (CLL) in an amount of 25 mg perday for 21 days followed by seven days rest on a 28 day cycle. Twentyseven patients of median age of 64 years (range; 47-75) were enrolled.Seventeen patients had Stage III or IV disease. Absolute lymphocytecounts were measured at Day 0, 7 and 30. Response was assessed at day 30and monthly thereafter using the NCI-WG 1996 criteria. Patients withstable disease or better response were continued on therapy for amaximum of 12 months while those with progressive disease receivedrituximab (375 mg/m²) added to Revlimid®. Patients were consideredevaluable for response if they completed at least two months oftreatment.

All patients were available for toxicity and 13 out of 18 patientsavailable for response evaluation. Nine patients on treatment were earlyfor response assessment. Five patients achieved complete response andfour patients achieved partial response. Three patients achieved stabledisease (continued on treatment). Overall response rate in the 13evaluable patients was 69%, while objective response rate defined as(complete response, partial response and stable disease) was 92.3%. Onlyone patient had progressive disease after three months of treatment.

Toxicity profile was predictable and manageable. Flare reaction (e.g.,tender swelling of lymph nodes, sinus congestion and/or rash) was thecommon side effects noted. Other side effects were tumor lysis syndrome,grade 3/4 hematologic toxicities, and febrile neutropenia.

The study result shows that Revlimid®is effective in treating leukemia,particularly chronic lymphocytic leukemia.

-   -   4-(Amino)-2-(2,6-dioxo(3-piperidyl))-isoindoline-1,3-dione        (Actimid™) was administered to patients with relapsed/refractory        multiple myeloma. The study was conducted in compliance with        Good Clinical Practices. Patients were at least 18 years old,        had been diagnosed with multiple myeloma (with paraprotein in        serum and/or urine), and were considered refractory to treatment        after at least two cycles of treatment, or have relapsed after        two cycles of treatment.

Patients who have progressive disease, according to the SouthwestOncology Group (SWOG) criteria, on their prior regimen are consideredtreatment refractory. Relapse following remission is defined as >25%increase in M component from baseline levels; reappearance of the Mparaprotein that had previously disappeared; or a definite increase inthe size and number of lytic bone lesions recognized on radiographs.Patients may have had prior therapy with thalidomide, provided they wereable to tolerate the treatment. A Zubrod performance status of 0 to 2 isrequired for all patients.

4-(Amino)-2-(2,6-dioxo(3-piperidyl))-isoindoline-1,3-dione isadministered to patients at doses of 1, 2, 5, or 10 mg/day for up tofour weeks; at each dose level, three patients are initially enrolled.Dosing occurs at approximately the same time each morning; all doses areadministered in the fasted state (no eating for at least two hours priorto dosing and two hours after dosing).4-(Amino)-2-(2,6-dioxo(3-piperidyl))-isoindoline-1,3-dione doses areadministered in an ascending fashion such that patients in the firstcohort received the lowest dose of4-(Amino)-2-(2,6-dioxo(3-piperidyl))-isoindoline-1,3-dione (1 mg/day)and escalation to the next higher dose level occurs only following theestablishment of safety and tolerability at the current dose. If one outof three patients at any dose level experience dose limiting toxicity(DLT), three additional patients are enrolled at that dose. If none ofthe three additional patients experience DLT, escalation to the nextdose level occurs; dose escalations continue in a similar fashion untilthe MTD is established or the maximum daily dose (10 mg/day) isattained. However, if one of the three additional patients enrolledexperiences DLT, the MTD has been reached. If two or more of the threeadditional patients enrolled experience DLT, the MTD is judged to havebeen exceeded and three additional patients are enrolled at thepreceding dose level to confirm the MTD. Once the MTD has beenidentified, four additional patients are enrolled at the dose level sothat a total of 10 patients is treated at the MTD.

Blood sampling for analysis of pharmacokinetic parameters is performedon Days 1 and 28 according to the following sampling schedule: pre-dose0.25, 0.5, 0.75, 1, 1.5, 2, 2.5, 3, 4, 6, 8, 10, 12, 18, and 24 hourspost-dose. An additional blood sample is collected at each weekly visitfor the determination of4-(Amino)-2-(2,6-dioxo(3-piperidyl))-isoindoline-1,3-dione levels. Totalurine collections are also made with urine pooled according to thefollowing time intervals post-dose: 0 to 4, 4 to 8, 8 to 12, and 12 to24 hours. Safety assessments are made by monitoring adverse events,vital signs, ECGs, clinical laboratory evaluations (blood chemistry,hematology, lymphocyte phenotyping, and urinalysis), and physicalexamination at specific times during the study.

Results of interim pharmacokinetic analyses obtained follwoing single-and multiple-dose administration of4-(Amino)-2-(2,6-dioxo(3-piperidyl))-isoindoline-1,3-dione to multiplemyeloma patients are presented below in Tables 1 and 2. These data showthat 4-(Amino)-2-(2,6-dioxo(3-piperidyl))-isoindoline-1,3-dione wassteadily absorbed at all dose levels in relapsed multiple myelomapatients. Maximum plasma concentrations occurred at a median T_(max) ofbetween 2.5 and 2.8 hours post-dose at Day 1 and between 3 and 4 hourspost-dose at Week 4. At all doses, plasma concentrations declined in amonophasic manner after reaching C_(max). The start of the eliminationphase occurred between 3 and 10 hours post-dose at Day 1 and Week 4,respectively.

These data also showed that after 4 weeks of dosing,4-(Amino)-2-(2,6-dioxo(3-piperidyl))-isoindoline-1,3-dione accumulatedto a small extent (mean accumulation ratios ˜1.02 to 1.52 and ˜0.94 to1.62 for C_(max) and AUC_((0−r)), respectively). There was almost a doseproportional increase in AUC_((0−r)) and C_(max) values with increasingdose. A five-fold higher dose of4-(Amino)-2-(2,6-dioxo(3-piperidyl))-isoindoline-1,3-dione produced a3.2- and 2.2-fold increase in C_(max) at Day 1 and Week 4, respectively.Similarly, a 5-fold increase in dose resulted in a 3.6- and 2.3-foldincrease in AUC_((0−r)), at Day 1 and Week 4, respectively.

TABLE 1 Pharmacokinetic parameters of Actimid ™ in relapsed multiplemyeloma patients 1 mg 2 mg 5 mg Parameter (N = 6) (N = 2) (N = 3) Day 1C_(max) ng/mL 15.03 (4.04) 24.4* (12.1) 48.56 (14.03) t_(max) h 3.3(2.6) 2.7* (0.3) 2.3 (0.3) AUC_((0-∞)) ng · h/mL 152.90 (36.62) 279.18(51.10) 593.10 (335.23) AUC_((0-τ)) 134.21 (27.14) 249.57 (29.26) 520.94(267.32) t½ h 7.3 (3.4) 6.3 (1.4) 6.5 (2.2) CL/F mL/min 114.75 (29.20)121.43 (22.22) 182.31 (117.06) Vz/f L 69.55 (44.97) 65.31 (2.80) 87.24(22.61) τ = 24 hours N/A = not available

TABLE 2 Pharmacokinetic parameters of Actimid ™ following multiple oraldoses(1, 2, and 5 mg/day) in relapsed multiple myeloma patients 1 mg 2mg 5 mg Parameter (N = 5) (N = 2) (N = 3) Week 4 C_(max) ng/mL 23.20(7.48)  30.05* (15.64)  58.07 (38.08) t_(max) h 3.6 (1.5) 2.8* (0.3) 5.0 (2.6) AUC_((0-∞)) ng · h/mL N/A N/A N/A AUC_((0-τ)) 239.31 (122.59)269.36 (186.34) 597.24 (354.23) t½ h 6.2* (0.6)  7.7 (2.8) 7.8 (4.0)CL/F mL/min 87.85 (48.48) 162.68 (112.54) 207.50 (175.41) Vz/f L 41.35*(8.84)  95.04 (35.39) 103.95 (27.25) τ = 24 hours N/A = not available *N= 3 patients

6.2.3 Treatment of Relapsed Multiple Myeloma

Two Phase 1 clinical studies of3-(4-amino-1-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione(Revlimid®) have been conducted to identify the maximum tolerated dose(MTD) in patients with refractory or relapsed multiple myeloma. Thesestudies have also characterized the safety profile of3-(4-amino-1-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione whenascending doses of3-(4-amino-1-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione weregiven orally for up to 4 weeks. Patients started3-(4-amino-1-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dionetreatment at 5 mg/day with subsequent escalation to 10, 25, and 50mg/day. Patients were enrolled for 28 days at their assigned dose, withthe option of extended treatment for those who did not exhibit diseaseprogression or experience dose limiting toxicity (DLT). Patients wereevaluated for adverse events at each visit and the severity of theseevents was graded according to the National Cancer Institute (NCI)Common Toxicity Criteria. Patients were discontinued if they experiencedDLT (Grade 3 or greater non-hematological, or Grade 4 hematologicaltoxicity).

In this study, 27 patients were enrolled. All patients had relapsedmultiple myeloma and 18 (72%) were refractory to salvage therapy. Amongthese patients, 15 had undergone prior autologous stem celltransplantation and 16 patients had received prior thalidomidetreatment. The median number of prior regimens was 3 (range 2 to 6).

Blood and urine samples were collected for analysis of pharmacokineticparameters on Days 1 and 28. Blood samples were collected according tothe following sampling schedule: pre-dose, 0.25, 0.5, 0.5, 0.75, 1, 1.5,2, 2.5, 3, 4, 6, 8, 10, 12, 18, and 24 hours post-dose. In addition ablood sample was collected at each weekly clinic visit for3-(4-amino-1-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dionedetermination. Total urine was collected and pooled according to thefollowing time intervals post-dose: 0 to 4, 4 to 8, 8 to 12, and 12 to24 hours. Response to treatment was assessed by M-protein quantification(by immunoelectrophoresis) from serum and a 24-hour urine collection,with creatine clearance and 24-hour protein calculations undertaken atscreening, baseline, Weeks 2 and 4, and monthly thereafter (or uponearly termination). Bone marrow aspirations and/or tissue biopsy arealso performed at Months 3, 6 and 12 if a patient's paraprotein serumconcentration or 24-hour urine protein excretion declined to the nextlower level, based on best response criteria. Preliminary results forthe 28-day treatment period are summarized below.

Preliminary pharmacokinetic analyses based on these two studiesindicated that AUC and C_(max) values increase proportionally with dosefollowing single and multiple doses in multiple myeloma patients (as wasseen in healthy volunteers). Further, there was no evidence ofaccumulation with multiple dosing as single dose AUC_((0−r)) wascomparable to multiple dose AUC_(0−r) following the same dose of3-(4-amino-1-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione.Similar to healthy volunteer studies, double peaks were observed.Exposure in multiple myeloma patients appeared to be slightly higherbased on C_(max) and AUC values as compared to healthy male volunteerswhile clearance in multiple myeloma patients was lower than it was inhealthy volunteers, consistent with their poorer renal function (both asa consequence of their age and their disease). Finally,3-(4-amino-1-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dionehalf-live in patients was shorter than in healthy volunteers (mean 8hours, ranging up to 17 hours).

In this study, the first cohort of 3 patients was treated for 28 days at5 mg/day without any dose limiting toxicity (DLT). The second cohort of3 patients subsequently commenced therapy at 10 mg/day. Patients in thesecond 10 mg/day of3-(4-amino-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione cohorttolerate treatment well.

-   -   6.2.4 Treatment of Relapsed or Refractory Multiple Myeloma

Patients with relapsed and refractory Dune-Salmon stage III multiplemyeloma, who have either failed at least three previous regimens orpresented with poor performance status, neutropenia or thrombocytopenia,are treated with up to four cycles of combination of melphalan (50 mgintravenously), an immunomodulatory compound of the invention (about 1to 150 mg orally daily), and dexamethasone (40 mg/day orally on day 1 to4) every four to six weeks. Maintenance treatment consisting of dailyand immunomodulatory compound of the invention and monthly dexamethasoneare continued until the disease progression. The therapy using animmunomodulatory compound of the invention in combination with melphalanand dexamethasone is highly active and generally tolerated in heavilypretreated multiple myeloma patients whose prognosis is otherwise poor.

The embodiments of the invention described above are intended to bemerely exemplary, and those skilled in the art will recognize, or willbe able to ascertain using no more than routine experimentation,numerous equivalents of specific compounds, materials, and procedures.All such equivalents are considered to be within the scope of theinvention and are encompassed by the appended claims.

1. A method of treating leukemia in a human, which comprisesadministering to a human in need thereof a therapeutically effectiveamount of3-(4-amino-1-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione.
 2. Themethod of claim 1, wherein the leukemia is chronic lymphocytic leukemia,chronic myelocytic leukemia, acute lymphoblastic leukemia, acutemyelogenous leukemia or acute myeloblastic leukemia.
 3. The method ofclaim 1, wherein the leukemia is relapsed, refractory or resistant toconventional therapy.
 4. The method of claim 1, wherein the leukemia ischronic lymphocytic leukemia.
 5. The method of claim 4, wherein theleukemia is refractory or relapsed chronic lymphocytic leukemia.
 6. Amethod of treating leukemia, which comprises administering to a patientin need thereof a therapeutically effective amount of3-(4-amino-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione and atherapeutically effective amount of a second active agent.
 7. The methodof claim 6, wherein the second active agent is antibody, hematopoieticgrowth factor, cytokine, anti-cancer agent, antibiotic, cox-2 inhibitor,immunomodulatory agent, immunosuppressive agent, corticosteroid, or apharmacologically active mutant or derivative thereof.
 8. The method ofclaim 6, wherein the second active agent is rituximab.
 9. The method ofclaim 8, wherein the leukemia is chronic lymphocytic leukemia, chronicmyelocytic leukemia, acute lymphoblastic leukemia, acute myelogenousleukemia or acute myeloblastic leukemia.
 10. The method of claim 8,wherein the leukemia is chronic lymphocytic leukemia.
 11. The method ofclaim 1, wherein the amount of3-(4-amino-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dioneadministered is from about 1 to about 50 mg per day.
 12. The method ofclaim 11, wherein the amount of3-(4-amino-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dioneadministered is about 5, 10 or 25 mg per day.
 13. The method of claim 1,wherein the amount of3-(4-amino-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dioneadministered is about 1, 2.5, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30,35, 40, 45, or 50 mg per day.
 14. The method of claim 1, wherein3-(4-amino-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dioneadministered is enantiomerically pure.
 15. The method of claim 14,wherein 3-(4-amino-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dioneadministered is S enantiomer.
 16. The method of claim 14, wherein3-(4-amino-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dioneadministered is R enantiomer.
 17. The method of claim 1, wherein3-(4-amino-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione isadministered in the form of a capsule or tablet.
 18. The method of claim1, wherein3-(4-amino-oxo-1,3-dihydro-isoindol-2-yl)-piperidine-2,6-dione isadministered in an amount of about 25 mg per day for 21 days followed byseven days rest in a 28 day cycle.
 19. The method of claim 18, furthercomprising administration of rituximab in an amount of 375 mg/m².